PROJECT SUMMARY/ABSTRACT Plasma membrane electron transport (PMET) is a ubiquitous system which oxidizes reducing equivalents (NADH and NADPH) and exports protons and electrons from the cell via Coenzyme Q10 (CoQ10). This pathway is increasingly recognized to be critical to normal cell function, and has not been characterized in islets. NADPH is a newly recognized metabolic coupling factor important for insulin secretion, although its mechanism of action has not been characterized. Our Preliminary Data demonstrate that pancreatic islets have substantial PMET activity, and we propose that the PMET is the primary sensor for the glucosedependent increase in cytosolic NAD(P)H and the signaling cascade leading to the secretion of insulin. To test this hypothesis, we will employ over-expression or knockdown using siRNA and adenoviral vectors strategies to analyze the role of components of the PMET pathway as the potential downstream targets in glucose metabolism and insulin secretion. We will analyze the role of NQO1, an oxidoreductase known to participate in the PMET pathway on the response of islets to glucose or other stimulatory agents. We will examine the role of components of the PMET system in glucose-stimulated insulin secretion (GSIS) and analyze the consequence of PMET pathway activation, namely PMET-dependent ROI production, on islet metabolism and kinase cascades, using a custom-made integrated electrochemical-confocal imaging platform to simultaneously study islet metabolism, respiration, and signaling. These studies will provide greater insight into the mechanism of NADPH-dependent insulin secretion and identify novel targets for the development of therapeutics for type 2 diabetes.